Effects of sodium butyrate on expression of members of the ICF-binding protein superfamily in human mammary epithelial cells

Dietary factors play an important role ill both the development and prevent ion of human cancers, including breast carcinoma. One dietary micronutrient , sodium butyrate (NaB), is a major end product of dietary starch and fiber , produced naturally during digestion by anaerobic bacteria ill the cecum a nd colon. NaB is a potent growth inhibitor and initiates cell differentiati on for many cell types in vitro. In this study, we investigated the effects of NaB on three human mammary epithelial cells and regulation of the IGF a xis, specifically, IGF-binding protein-3 (IGFBP-3), a known growth regulato r ill human mammary cells, and IGFBP-related protein 2 (IGFBP-rP2)/connecti ve tissue growth factor. NaB inhibited DNA synthesis, as measured by [H-3]thymidine incorporation, i ll estrogen-responsive (MCF-7) and estrogen-non-responsive (Hs578T) breast cancer cells, and normal human mammary epithelial cells (HMEC) to a similar degree (up to 90% inhibition at 1-10mM concentrations). Treatment of cells with NaB induced histone hyperacetylation, suggesting that NaB exerts its biological effects, at least in part, as a histone deacetylase inhibitor in mammary epithelial cells. Treatment of Hs578T cells with NaB caused an ind uction of apoptotic cell death. NaB treatment resulted in increased levels of p2(1Waf1/Cip1) mRNA and protein in Hs578T cells and distinct upregulatio n of p27(Kip1) in HMEC, suggesting that NaB activates different genes invol ved in cell cycle arrest, depending upon the cell type. In the same context , among the IGFBP superfamily members tested, NaB specifically upregulated the expression of IGFBP-3 and IGFBP-rP2. These two proteins are known to be involved in inhibition of mammary epithelial cell, replication. Northern b lot analysis showed that NaB treatment at 1-10mM concentrations caused a do se-dependent stimulation of IGFBP-3 mRNA expression in cancerous cells and IGFBP-rP2 mRNA expression in both cancerous and non-cancerous cells. Protei n data from Western ligand blot and immunoblot analyses demonstrated parall el results. In summary, we have demonstrated that NaB (i) uniformly suppresses DNA synt hesis in both cancerous and non-cancerous mammary cells, and (ii) upregulat es IGFBP-3 and IGFBP-rP2 mRNA and protein levels in cancerous and non-cance rous mammary cells. These results provide the first demonstration that buty rate regulates the IGFBP system in the human mammary system.